Mammalian hemoglobins are composed of two pairs of chemically distinct subunits, designated alpha and beta in the case of adult human hemoglobin. The association of the alpha and beta chains results in a tetrameric molecule which is capable of cooperative ligand binding and, therefore, efficient oxygen transport. In addition, deoxyhemoglobin exhibits a higher affinity for organic phosphates and protons than the liganded form, and these properties result in a linkage between oxygen transport, carbon dioxide transport, and erythrocyte metabolism. Since the primary structures of the two types of subunits are different, the alpha and beta chains are neither expected nor observed to make an equal contribution to the physiological properties of hemoglobin. The structural causes of these differences in reactivities of the chains will be investigated by varying the size of the heme ligand, by examining the ligand binding properties of minor hemoglobin components and hemoglogins from different species, and by substituting unnatural hemes for protoheme in human hemoglobin A. Studies of the differential release of anions and protons associated with ligand binding to the individual chains are also proposed. Finally, the reaction of the isolated alpha and beta chains to form native tetrameric hemoglobin will be examined. The majority of the proposed experiments involve kinetic measurements using rapid mixing stopped-flow techniques. The studies are designed to provide new information about the relative importance of the alpha and beta chains in determining the physiological properties of hemoglogin. Information of this type, especially in terms of the effects of specific amino acid modifications, is valuable for the treatment and interpretation of clinical symptons produced by hereditary hemoglobinopathies (viz. hemoglobin S, C, I, M, etc.) and related medical problems concerning impaired oxygen transport and cardiovascular functions (viz. thalassemia, hypoxia, polycythemia, etc.). Similarly, studies of the individual properties of the chains are required for an understanding of the evolutionary development of hemoglobin and of the importance and function of the various minor hemoglobin components.